1. Field of the Invention
This invention relates to apparatus and methods for liquid-solid column centrifugation chromatography including automated, continuous systems therefor.
2. Description of the Prior Art
Chromatography is an analytical and preparative chemical process which separates molecules from each other according to their various physical and chemical properties, including their molecular size, electric charge and tendency to adsorb to various specific surfaces. This method is widely used throughout the chemical and medical sciences in research work, clinical laboratories and industry, both to analyze mixtures of chemicals and to purify specific chemcals. Liquid-solid chromatography involves the chromatography of molecules dissolved in liquids which are passed over beds of particles. When the particles are contained in a column, a porous plate is generally placed at the bottom and the particle bed is packed thereover. The porous plate serves to support the particle bed while allowing liquid to flow through the column and bed.
Among the problems which exist with known types of liquid-solid chromatography, such as gel filtration or permeation chromatography is the fact that the technique is frequently unduly time consuming. Also, the process frequently leads to undesirable dilution of the molecules being chromatographed.
It has previously been suggested to employ the centrifuge in chromatography.
It has been suggested to employ centrifugal gel filtration methods in connection with basket centrifuges. See "A Procedure for Gel Filtration of Viscous Solutions" by N. I. Arne Emneus, 32 Journal of Chromatography, pages 243-257 (1968). Other disclosures of the use of a centrifuge in chromatography as well as tube construction and methods are found in "A Rapid Method for Desalting Small Volumns of Solution" by Neal and Florini, 55 Analytical Biochemistry, pages 328-330 (1973) and "Micro-Step-Exclusion Chromatography" by Krieger and Tobler, 81 Analytical Biochemistry, pages 450-453 (1977).
U.S. Pat. No. 3,583,230 discloses sample injection methods and apparatus. The injector has an inlet opening which communicates with a central channel provided with chromatographic filter material and a capillary outlet. It is noted, however, the injection of the liquid samples into the column is for subsequent chromatographic development without use of centrifugation.
U.S. Pat. No. 3,440,864 discloses liquid chromatographic columns with a threaded jacket. The system is not designed for centrifugal chromatography.
U.S. Pat. No. 3,664,845 discloses the use of a bed of molecular fractionating gel under the influence of a centrifugal force such that the bed is expanded and contracted by means of pulsed centrifugation.
It has been known to attempt various means for automating chromatography. U.S. Pat. No. 3,194,400 relates to a liquid chromatographic centrifuge wherein a rotor cooperates with radial channels which are provided with separate media. The disc 20 is provided with radial channels 40. After centrifugation, the various zones are examined to determine different zones of chemical fractions. These zones may then be separately eluted from portions of the sheet. The receptacles provided are intended to receive solvent liquid flowing from the sheet, but generally this liquid would not be collected in discrete fractions. The resolved chemicals stay on the sheet.
While not involving centrifugation chromatography in columns, U.S. Pat. No. 3,395,093 discloses separations which are carried out on a disc of paper or gel under the dual influence of centrifugal and electrical forces. Structure for handling the materials is disclosed.
U.S. Pat. No. 3,666,105 discloses continuous liquid-solid chromatography employing a pair of cylinders which cooperate to define a column. It is noted that the fraction collector rotates with the column as a result of the design.
A number of other disclosures suggest the use of discs. U.S. Pat. No. 4,077,886 employs discs and makes no provision for collection of the resolved components into eluted fractions for analysis and subsequent use. U.S. Pat. No. 3,417,548 discloses the use of a pair of spaced discs. U.S. Pat. No. 3,527,350 discloses centrally introduced flow and resolution within a disc bed. U.S. Pat. No. 3,617,557 discloses continuous centrifugation chromatrography in a disc material. Under the influence of Coriolis forces, different separated components come to different regions of the rim of the disc. Cups are provided to receive eluate from discrete regions of the rim. U.S. Pat. No. 3,201,921 discloses selective treatment of a fluid by adsorption. Pumping action is the primary moving force for the liquid which is fed in at least two zones with a plurality of cells being provided adjacent the periphery. U.S. Pat. No. 3,113,103 discloses the use of centrifugal force in the disc. No provision is made for elution of separated components and, as a result, no fraction collector is provided. Means are provided for loading of samples.
There remains, therefore, a significant need for effective apparatus for both batch and continuous liquid-solid column centrifugation chromatography which produces improved resolution in reduced time.